Numerical source data.

Ranaviruses are capable of infecting both wild and farmed fish, amphibians, and reptiles, leading to significant economic losses and ecological risks. Currently, ranaviruses have been found in at least 175 species spanning six continents. Except for Singapore grouper iridovirus (SGIV), ranavirus genomes are generally regarded as highly methylated. Nevertheless, our comprehension of the methylation characteristics within ranaviruses remains limited. Despite the numerous genomes currently included in the GenBank database, a complete phylogenetic tree for ranaviruses has not yet been determined, and interspecific evolutionary relationships among ranaviruses have not been thoroughly investigated. In this study, the whole-genome methylation profile of mandarin fish ranavirus (MRV; a ranavirus) was investigated, revealing a methylation level of 16.04%, and hypomethylation of the MRV genome was detrimental to viral replication, speculating the genome methylation may play an important role in MRV replication. Furthermore, by combining with whole-genome DNA sequence phylogenetic analyses, we propose the possibility of an interspecies evolutionary relationship among ranaviruses, with the presence of four distinct evolutionary lineages within ranavirus evolution: "SGIV, SCRAV(MRV/LMBV), EHNV/ENARV/ATV, and CMTV/FV3", which might be also supported by the genomic collinearity, natural host range and host habitats. Furthermore, ranavirus genomic methylation levels may provide additional evidence for this hypothesis, but further proof is needed. Our work enhances the understanding of the role of genome methylation in ranaviruses and is beneficial for the prevention and control of ranavirus diseases; simultaneously, the proposed evolutionary hypothesis of ranavirus provides novel insights and ideas for exploring the evolutionary trajectory of viruses.